2-Chlorodeoxyadenosine (CdA) is an adenosine deaminase-resistant congener of deoxyadenosine with selective toxicity towards lymphocytes and monocytes. A clinical trial of CdA therapy for rheumatoid arthritis (RA) is in progress at our institution and at the Clinical Research Center of a collaborating local medical center. The goal of the proposed Research plan is to characterize monocyte phenotype and function changes in RA patients during CdA therapy, and to further elucidate the mechanism of CdA action by pertinent in vitro biochemical assays. Preliminary results show a complete disappearance of circulating monocytes within five days in RA patients receiving CdA by continuous infusion. Furthermore, sub-toxic concentrations of CdA in vitro inhibit monocyte phagocytosis and the secretion of the inflammation mediator interleukin-6 (IL-6). The mechanism by which CdA exerts anti-monocyte effects is unknown, and may involve DNA cleavage by an endonuclease (programmed cell death). Also, differential effects of CdA towards monocyte/macrophage subpopulations, at different stages of maturation or activation, have not been fully explored. The specific aims of the proposed research are (1) to further examine the metabolism, function, and DNA integrity of normal donor monocytes exposed in vitro to CdA; (2) to measure sequential changes in monocyte phenotype, DNA fragmentation pattern, and plasma cytokine levels in RA patients receiving CdA by intermittent infusion; and (3) to correlate surface marker and cytokine changes with clinical outcome as determined by detailed joint assessment and performance index. Surface marker studies on cultured or RA patient monocytes will be performed by flow cytometry using monoclonal antibodies against differentiation antigens CD33 and CD14, and against class II MHC antigens HLA-DR and DQ. Plasma or culture supernatant levels of IL-1, tumor necrosis factor, and soluble IL-2 receptors will be measured by ELISA, and IL-6 levels will be determined by bioassay. In addition, urinary levels of neopterin will be monitored by RIA and HPLC. Gel electrophoresis of monocyte DNA will be used to detect the internucleosomal cleavage pattern typical of apoptosis. Together, these studies will determine important biochemical and immunological characteristics of CdA toxicity towards human monocytes, and will form the basis for designing better treatment strategies in RA using CdA and other agents that affect mononuclear phagocytes.